Surface hybridization studies

As it was discussed in detail before, the whole association / dissociation process can be described by a simple Langmuir model with the two rate constants, 1 and koffi respectively, describing the complete process. This is indicated in Figme 12 by the full curve through the experimental data points. The agreement is excellent and the obtained rate constants are given in Table II. 

This experiment is a good example for a very specific feature of SPFS being a label-detecting senior scheme it still does not necessarily mean that the analyte has to be labeled. The surface binding reaction-induced change of chromojduKe prqierties, in this case the chromophore / metal separation distance, allows fw a sensitive detection of an analyte without the need to have it chemically modified. 

PROTEDJ BINDING STUDIES - THE LIMIT OF DETECTION IN SPFS 

Of course, there is no reason to limit the SPFS detection schemes to surface hybridization studies labeled proteins can be detected as sensitively. 

Rather than giving a number of additional examples for the use of SPFS in proteomics we will briefly discuss only one particularly important aspect, i.e., the limit of detection (LOD) in surface binding studies. 

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We then introduce a few examples for the use of SPFS, first in surface hybridization studies and then for antigen-antibody interaction assays. We will give, in particular, examples for different versions of fluorescence spectroscopy making use of, e.g., donor-acceptor energy transfer phenomena between correspondingly labeled binding partners. 

Figure 11. Various sur ice architectures and experimental schemes for surface hybridization studies (a) a chromophore-Iabeled analyte hybridizes to an unlabeled sur ce-attached probe oligonucleotide strand (b) a chromophone-labeled, sur ce-ottached probe strand stretches upon hybridizing to an unlabeled target analyte strand, thus dacii the chtomophore fi er away (by Ad) from the quenching metal sur e (c) scheme firr a fluoiescence resonance energy tiansfer (FRET) experiment between a donor-dye labeled sur ce-attached probe oligonucleotide strand and an iuxteptor-dye labeled analyte target strand hybridized from solution.

The last example fw SPFS-based surface-hybridization studies is schematically depicted in Figure 11(c) Here, the probe strand is labeled with a chromophore that acts as a donor dye for an acceptor chromophore chemically linked to the target strand. Before hybridization the angular scan of the fluorescence shows the typical features of a SP-... 

A multi-layer surface architecture composed of SAM/streptavidin/probe was employed for the hybridization study (Fig. 19). Since the streptavidin density on the functional stripes was identical to that on a homogenous surface [9], i.e., 2.2 x 1012 molecules cm-2, the probe density was estimated to be 2.9 x 1012 molecules cm 2 by knowing from the diffraction signal the binding stoichiometry between streptavidin and the probe (ca. 0.75) [16],... 

We recently considered the effect of the nucleic acid-surface electrostatic interaction on the thermodynamics of the surface hybridization [2-5, 22], This theory used an analytical solution of the linearized Poisson-Boltzmann boundary value problem for a charged sphere-surface interaction in electrolyte solution and corresponds to the system characterized by a low surface density of immobilized probes. To understand the motivation for that work and extensions, we need to consider the physical effects of a surface in solution and the theoretical tools available for their study. 

Present study deals with specific aspects of development of hybrid proteins capable to create p-sheeted protein fibrils and integration of the fibrils with solid surfaces. Our study aims to define the most essential conditions and parameters on which the arrangement of the complex object on the solids depends. Morphology,... 

In the following, a few examples for the use of surface plasmon fluorescence spectroscopy in DNA hybridization studies will be given. The general architecture of the sensor surface layer has already been introduced and is summarized in Figure 11. 

Stioppa A, Kresse G (2008) The shortcomings of semi-local and hybrid functionals what we can learn frinn surface science studies. New J Phys 10(6) 063020... 

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